Koonin E V, Agol V I
Virology. 1983 Sep;129(2):309-18. doi: 10.1016/0042-6822(83)90170-8.
Replication complexes (RC) containing endogenous viral RNA templates and the viral RNA polymerase were isolated from encephalomyocarditis (EMC) virus-infected Krebs II cells by two different procedures one of which included removal of the bulk of the associated proteins by treatment with 2 M LiCl. Replacement of one or all the four nucleoside triphosphate (NTP) substrates by corresponding nucleoside diphosphates (NDPs) did not eliminate the ability of the RC to synthesize viral RNA products. Moreover, the complexes were shown to accept as substrates even nucleoside monophosphates (NMPs), provided at least one NTP was present in the system. These results suggested that nucleotide kinases, NMP kinase and NDP kinase, were associated with the RC, and this suggestion was confirmed directly. The RC could be resolved, by sucrose gradient centrifugation, into distinct components that exhibited marked preference for either NDPs or NTPs as substrates for RNA synthesis. It is suggested that, in the NDP-preferring components, the NTP molecules generated by the built-in nucleotide kinase system are channeled directly to the vicinity of the replication fork.
通过两种不同的方法，从感染脑心肌炎（EMC）病毒的克雷布斯II细胞中分离出含有内源性病毒RNA模板和病毒RNA聚合酶的复制复合物（RC），其中一种方法包括用2M LiCl处理以去除大部分相关蛋白质。用相应的核苷二磷酸（NDP）取代四种核苷三磷酸（NTP）底物中的一种或全部，并不会消除RC合成病毒RNA产物的能力。此外，即使系统中至少存在一种NTP，这些复合物也被证明能够接受核苷单磷酸（NMP）作为底物。这些结果表明，核苷酸激酶，即NMP激酶和NDP激酶，与RC相关，这一推测得到了直接证实。通过蔗糖梯度离心，RC可以分解为不同的组分，这些组分在作为RNA合成底物时，对NDP或NTP表现出明显的偏好。有人提出，在偏好NDP的组分中，由内置核苷酸激酶系统产生的NTP分子会直接被引导到复制叉附近。
